The central hypothesis of this proposal is that there are chronic lymphocytic leukemia (CLL) associated antigens that can be recognized by host T cells. However, due to the CLL phenotype the T cells that can recognize such antigens may be rendered anergic. Conventional and some novel forms of chemotherapy for this disease may adversely affect anergic T cell that may be capable of responding to CLL-associated antigens. CLL cells can be converted into effective antigen presenting cells via CD40-ligation in vitro. This led to development of strategies for introducing the gene encoding the CD40-ligand (CD154) into CLL cells using recombinant adenovirus vectors, designated AdCD154. We find that CLL cells infected with AdCD154 can induce autologous T cell responses in vitro, leading to generation of cytotoxic T lymphocytes (CTL) against non-modified CLL cells. This served as the basis for a phase I gene therapy trial intended to examine the safety of a single intravenous infusion of autologous AdCD154-CLL cells to patients with intermediate or high-risk disease. The encouraging results from this clinical trial and preliminary in vitro studies support the hypothesis that there are CLL-associated antigens that can be recognized to host T cells and potentially targeted by host CTL, leading to immune clearance of CLL cells. To dissect the spontaneous and induced cellular responses to putative CLL-associated antigens we have the following specific aims. (1) Examine the expressed T cell receptor Vbeta repertoires of blood T cells of patients at diagnosis and pre- and post treatment with therapies developed by the CLL Research Consortium (CRC). (2) Evaluate the precursor frequency of blood T cells at similar time-points that can elaborate interferon-gamma in response to AdCD154-CLL or to autologous CLL cells after CD3/CD28-ligation in vitro. (3) Define peptides encoded by genes implicated in the pathogenesis or progression of CLL by in Project 1 of this proposal that could serve as potential LL- associated antigens. (4) Use T cell lines reactive with autologous CLL cells in strategies that can identify and isolate genes encoding tumor- associated antigens. (5) Develop a phase II CRC trial of gene therapy involving multiple infusion of autologous AdCD154-CLL cells. Through these studies we identify CLL-associated antigens and develop novel forms of immune-based therapies for this disease.